Method of stimulating the immune response with halogenated 10-(ω-dialkylaminopolymethyleneamino)-2-methoxypyrido[3,2-b]quinolines

ABSTRACT

A method of stimulating the immune response in warm-blooded animals which comprises the administration of a halogenated 10-(ω-dialkylaminopolymethyleneamino)-2-methoxypyrido [3,2-b]quinoline.

BACKGROUND OF THE INVENTION

The compound7-chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolineis disclosed in Chemical Abstracts, 45, 8531b (1951) and Journal of theChemical Society, 2448-2455 (1954) as an antimalarial agent. It iscommercially available as the dihydrochloride salt from Aldrich ChemicalCompany.

SUMMARY OF THE INVENTION

This invention is concerned with a method of stimulating the immuneresponse in a warm-blooded animal which comprises internallyadministering to said animal an effective immunostimulating amount of acompound of the formula: ##STR1## wherein R₁ and R₂ are each selectedfrom the group consisting of hydrogen and halogen; A is a straight orbranched alkyl chain of 3 to 7 carbon atoms; B is selected from thegroup consisting of dimethyl, diethyl and di-(2-hydroxyethyl); and thepharmaceutically acceptable salts thereof.

DESCRIPTION OF THE INVENTION

The compounds of the present invention may be prepared according to thefollowing reaction sequence. ##STR2##

In accordance with this sequence a10-chloro-2-methoxypyrido[3,2-b]quinoline (1) is reacted with phenol anda dialkylaminoalkylamine with heat to give compound (2) which is thendiluted with ethanol and poured into a mixture of acetone andhydrochloric acid to produce the dihydrochloride.

Among the compounds contemplated by the present invention are thefollowing:

7-Chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinoline dihydrochloride,

7-Chloro-10-[(4-diethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride,7-Chloro-10-[(4-dimethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride,

7-Chloro-10-[(7-diethylaminoheptyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride,

7-Chloro-10-[(6-dimethylaminohexyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride,

8-Chloro-10-[4-(dimethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride,

7-Chloro-10-[(5-diethylaminopentyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride, and

2,2'-[[3-[(7-Chloro-2-methoxypyrido[3,2-b]quinolin-10-yl)amino]propyl]imino]diethanoldihydrochloride.

Immunotherapy is a new therapeutic approach to the treatment of cancerand is based on the concept that there are distinctive antigens in or onmost tumor cells that distinguish them from normal host cells. Amajority of tumor immunologists favor the view that potentiallymalignant cells constantly arise but because of their "foreigness" theyare normally eliminated by a competent humoral and cellular immunesystem. Occasionally, however, tumor cells escape this immunesurveillance and continue to reproduce--and cancer results. The reasonsfor the failure of the normally efficient immune surveillance mechanismsare not completely understood but it is thought that the immune systembecomes less effective with increasing age. It is depressed in certaingenetic immunodeficiency diseases, in various bacterial, fungal or viralinfections, and in patients undergoing immunosuppressive therapy. Thegrowth of the neoplasm itself, as well as the various therapeuticmodalities designed to treat the disease, e.g., cytotoxic chemotherapyand irradiation, leads to a still greater depression of host resistanceand results in an increased susceptibility to both exogenous andendogenous infections and perhaps accounts for the re-initiation oftumor growth and metastasis which all too frequently followstreatment-induced tumor remission.

If depression of the immune system facilitates the growth ofmalignancies, stimulation of immune responses may help the host toovercome residual cancer cells. Therefore it is considered desirable tosearch for chemical agents (i.e., immunostimulants) capable of restoringor stimulating the host's own immune defense mechanisms in order toovercome the deficiencies which account for the increased susceptibilityto disease and failure to eradicate the cancer. It is acknowledged thatsuch immunostimulating agents would very likely be incapable ofarresting the growth of a large, rapidly proliferating tumor but thattheir clinical utility would derive from their capacity to enhancenormal immune surveillance mechanisms in patients whose tumor burden hadlargely been reduced by conventional surgical, radiotherapeutic orchemotherapeutic methods. It would be hoped that the few remaining tumorcells could then be destroyed immunologically, producing a higherincidence of long term survivors or complete cures.

Experimental studies in animals have demonstrated the antitumorpotential of a number of immunostimulants including live organisms ofbacillus Calmette-Guerin (BCG), heat-killed cells of Corynebacteriumparvum, polynucleotides, and the anthelmintic drug, levamisole. Thesesubstances have been shown to stimulate cellular immunity and to producetumor regressions. Some successes have been claimed in early clinicaltrials with BCG against malignant melanoma and acute leukemia and withlevamisole against lung cancer and breast cancer. Although the antitumoreffects produced by these agents have been promising, significanttherapeutic benefits have yet to be realized. Since this is a very newtherapeutic approach, new drugs and methods of treatment must receivecareful clinical evaluation in order to reveal the full potential ofthese drugs.

Modern research is directed to the discovery of a drug similar to, butmore potent than, the known immunostimulants such as levamisole thatwould be effective in the eradication of tumor cells when used inconjunction with standard therapeutic measures. Stimulators of hostresistance may be detected in animal models that can, in fact, detectboth immunostimulators and anticancer agents. Mice are put in acondition simulating the immunodepression common to cancer patients.This is accomplished by infecting mice either with a leukemia viruswhich produces both leukemia and a disease-related immunodepression orwith a transplantable mammary tumor. Effective drugs are recognized bytheir ability to restore or enhance the antibody response in theexperimental mice.

Still another means of recognizing drug-induced stimulation of theimmune response is to measure increased antibody responses or increasedprotective effects produced by the coadministration of vaccines and"immunoadjuvants" such as the well-known effects of Freund's adjuvantand the more recently described effects of chemicals such as levamisole.See Brugmans, J., et al., Restoration of Host Defense Mechanisms in Manby Levamisole, Life Sciences 13: 1499-1504, 1973 and Renoux, G. andRenoux, M., Stimulation of Anti-Brucella Vaccination in Mice byTetramisole, a Phenyl-Imidothiazole Salt, Infect. & Immunity 8: 554-548,1973.

A further discussion of the function of immune response, methods ofstimulation and testing, may be found in the following references,Stimulation of Humoral and Cellular Antibody formation in Mice by PolyI:C, W. Turner, et al., Proc. Soc. Exp. Biol. & Med., 133, 334-338(1970) and Humoral and Cellular Immune Responses in Susceptible andResistant Strains of Mice Infected with Friend Leukemia Virus, W. S.Cezlowski, et al., Proc. Soc. Exp. Biol. & Med., 146, 619-624 (1974).

The compounds of the present invention are active as immunostimulantswhen tested according to the following procedures:

(A) Rauscher leukemia virus

Rauscher leukemia virus is inoculated intraperitoneally into BALB/Cmice. The virus inoculum is a 20% (W/V) spleen extract made from 21-dayinfected spleens of BALB/C mice. All mice are within a three gram weightrange, with a minimum weight of 18 g., and all mice are of the same sex,usually male. Sheep red blood cells are injected intraperitoneally onthe seventh day. There are 5 mice per test group. The test compound isadministered orally on the sixth day as 0.5 ml. (in 0.2% Noble agar insaline) at a dose of 37.5 to 600 mg/kg of body weight, and again on theseventh and eighth day, in the same manner. On the fourteenth day themice are weighed and bled from the retro-orbital sinus. The blood ispooled and the harvested serum is stored at 4° C. for 24 hours.Hemagglutinin tests are performed by standard procedures using themicrotiter plate technique. Acceptable hemagglutinin titer for leukemic(immunosuppressed) mice is ≦1:128. Positive control compounds are PolyI:C (polyinosinic acid:polycytidylic acid) administeredintraperitoneally on days +6, +7 and +8 and/or Tilorone given orally ondays +6, +7 and +8. Acceptable positive control hemaglutinin titers are4-fold higher than the titers obtained in the leukemic control mice.

The results of this test, using representative compounds of thisinvention, appear in Table I.

                                      TABLE I                                     __________________________________________________________________________    Effect of Treatment on Antibody Response to Sheep Red Blood Cells in          Leukemic Mice                                                                                   Dose                                                                              Hemmagglutinin Titer*                                   Compound          mg/kg.                                                                            Test 1                                                                             Test 2                                                                             Test 3                                                                             Test 4                                                                             Test 5                                                                             Test 6                         __________________________________________________________________________    7-Chloro-10-(4-diethylamino-1-                                                                  600      512                                                methylbutylamino)-2-methoxypyrido                                                               300 512  512  256  256  512  256                            [3,2-b]quinoline dihydrochloride                                                                200           128  128                                                        150      128            256  128                                              100           128  64                                                         75       64             64                                                    50                 128                                                        37.5     32                                                 Poly I:C          10  2048 1024 512  512  1024 512                            Tilorone          200 1024 512                                                Non-Infected, Immunized (sheep red                                            blood cell) Control                                                                             --  4096 2048 1024 1024 1024 1024                           Infected, Immunized Control**                                                                   --  64   32   64   64   128  64                             7-Chloro-10-[(4-diethylaminobutyl)                                                              300 64   256                                                amino]-2-methoxypyrido[3,2-b]quino-                                                             150 128  128                                                line dihydrochloride                                                                            75  128  128                                                Poly I:C          10  512  512                                                Non-Infected, Immunized (sheep red                                            blood cell) Control                                                                             --  1024 1024                                               Infected, Immunized Control                                                                     --  32   64                                                 7-Chloro-10-[(4-dimethylaminobutyl)                                                             300      512  256  128                                      amino]-2-methoxypyrido[3,2-b]quino-                                                             150 256  256  256  64                                       line dihydrochloride                                                                            75  64   64   128  64                                                         37.5                                                                              64                                                                        18.75                                                                             64                                                      Poly I:C          10  512  1024 512  512                                      Non-Infected, Immunized (sheep red                                            blood cell) Control                                                                             --  2048 2048 1024 512                                      Infected, Immunized Control                                                                     --  32   32   64   32                                       7-Chloro-10-[(7-diethylaminoheptyl)                                                             300 256  512                                                amino]-2-methoxypyrido[3,2-b]                                                                   150 64   64                                                 quinoline dihydrochloride                                                                       75  32                                                      Poly I:C          10  512  512                                                Non-Infected, Imminized (sheep red                                            blood cell) Control                                                                             --  1024 1024                                               Infected, Immunized Control                                                                     --  32   64                                                 7-Chloro-10-[(6-dimethylaminohexyl)                                                             300 128  128                                                amino]-2-methoxypyrido[3,2-b]                                                                   150 128  128                                                quinoline dihydrochloride                                                                       75  64   64                                                 Poly I:C          10  1024 512                                                Non-Infected, Immunized (sheep red                                            blood cell) Control                                                                             --  2048 1024                                               Infected, Immunized Control                                                                     --  32   32                                                 8-Chloro-10-[(4-dimethylaminobutyl)                                                             300 256  256                                                amino]-2-methoxypyrido[3,2-b]qui-                                                               150 64   256                                                noline dihydrochloride                                                                          75  128  128                                                Poly I:C          10  512  512                                                Non-Infected, Immunized (sheep red                                            blood cell) Control                                                                             --  1024 1024                                               Infected, Immunized Control                                                                     --  64   64                                                 __________________________________________________________________________     *Reciprocal of serum dilution producing at least 50% agglutination of         sheep red blood cells.                                                        **Mice infected 7 days prior to injection of sheep red blood cells with       Rauscher leukemia virus.                                                 

(B) Immunochemotherapy of Lewis Lung Carcinoma

The effect of immunostimulating compounds on the antitumor activity ofreference cytostatic agents was demonstrated in a model in which anineffective and toxic dose of cytoxan was given to mice bearing theLewis lung carcinoma followed by treatment with the immunostimulant.BDF₁ mice were inoculated intramuscularly with 1×10⁵ Lewis lung cellsand treated with a single IP dose of cytoxan at 200 mg/kg three dayslater. Treatment with the subject compounds and known immunostimulantscommenced 3 day following the cytoxan with single oral doses being givenat 4 day intervals for a total of 4 doses. Test compounds (potentialimmunostimulants) were considered active if they provided a significantincrease in long term survivors compared to the animals treated withcytoxan alone.

When7-chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride was administered by the oral route at 300 mg/kgfollowing cytoxan treatment, there was a highly significant increase inlong term survivors compared to the nontreated tumor group and thecytoxan-only group. The known immunomodulators pyran and poly I:C alsoproduced increased survival times when given in combination withcytoxan. The most striking effect of7-chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride and the known immunomodulators was their capacity toreduce the toxicity of cytoxan in Lewis lung-bearing mice. The resultsof this study are recorded in Table II.

                                      TABLE II                                    __________________________________________________________________________    Immunochemotherapy of Lewis Lung Tumor in BDF.sub.1 Mice                      First.sup.1                                                                        Second.sup.2                                                             Drug Drug   (mg/kg)                                                                            Route                                                                             MST  % ILS.sup.3                                                                         S/T                                           __________________________________________________________________________    Placebo                                                                            Placebo                                                                              Saline                                                                             IP  42.0 --    3/18                                          Cytoxan                                                                            Placebo                                                                              Saline                                                                             IP  14.0       0/12.sup.4                                    Cytoxan                                                                            Pyran  20.0 IP  >60.0                                                                              >329  11/12                                         Cytoxan                                                                            Poly I:C                                                                             10.0 IP  >60.0                                                                              >329  9/12                                          Cytoxan                                                                            7-Chloro-                                                                            300.0                                                                              Oral                                                                              >60.0                                                                              >329  11/12                                              10-[(4-                                                                       diethylam-                                                                    ino-1-meth-                                                                   ylbutyl)                                                                      amino]-2-                                                                     methoxy-                                                                      pyrido                                                                        [3,2-b]                                                                       quinoline                                                                     dihydro-                                                                      chloride                                                                 Cytoxan                                                                            7-Chloro-                                                                            150.0                                                                              Oral                                                                              31.0  121  3/12                                               10-[(4-                                                                       diethylam-                                                                    ino-1-meth-                                                                   ylbutyl)                                                                      amino]-2-                                                                     methoxy-                                                                      pyrido                                                                        [3,2-b]                                                                       quinoline                                                                     dihydro-                                                                      chloride                                                                 __________________________________________________________________________     .sup.1 Cytoxan administered to all groups, except Placebo controls, by        single IP injection on day 3 at 200 mg/kg.                                    .sup.2 Immune modulators administered once daily on days 6, 10, 13 and 17     3 Percent ILS relative to cytoxan + placebo treatment group.                   .sup.4 Control for all treatment groups that received cytoxan. Cytoxan       treatment at 200 mg/kg IP on day 3 resulted in premature death of             tumorbearing mice.                                                            MST = Median Survival Time in days.                                           % ILS = Percent Increase in Life Span.                                        S/T = Survivors/Total.                                                   

(C) Adjuvant Effect on Influenzavirus Vaccine

Swiss white mice were immunized subcutaneously with 0.5 ml of aninactivated, solvent-extracted influenzavirus vaccine (A/Port Chalmers)diluted to contain 30 CCA of virus antigen. Compounds to be tested asimmunoadjuvants were administered by gavage as single, daily doses onthe day before, the same day, and the day after vaccination.Experimental and control mice were infected 21 days followingvaccination by the intranasal instillation of 0.05 ml of a dilution ofvirulent influenzavirus (A/Port Chalmers) estimated to kill 50% of thenon-immunized, control mice. The influence of orally administeredimmunoadjuvants on the protective effects of influenza vaccine wasdetermined on the basis of differences in the extent of pulmonarydisease (pneumonia) occurring in the vaccine plus drug groups comparedto the vaccine-only groups.

Mice treated with7-chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride at the time of vaccination showed survival ratiossimilar to mice receiving the vaccine alone. However, among the micesurviving challenge with influenzavirus, those receiving7-chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride in addition to vaccine showed a lesser degree ofpulmonary disease (i.e., lower lung weight/body weight ratio) than themice receiving vaccine alone. These results are set forth in Table III.Similar studies showing potentiation of influenza vaccine withadditional compounds of this invention are shown in Tables IV and V.

                                      TABLE III                                   __________________________________________________________________________    "Adjuvant" Effect on the Immunogenicity of Influenzavirus A Vaccine           (Pooled Data from 3 Experiments)                                                          Day of Treatment                                                   Treatment Group                                                                           Relative to Infection Vaccine.sup.1 Drug.sup.2                                           Survivors/Total (14 Days Post-Infection)                                                   ##STR3##                                 __________________________________________________________________________    Vaccine Control                                                               21               None   27/30       1.24                                      7-Chloro-10-[(4-                                                              21                                                                            22-21-20    29/30                                                                               1.03.sup.3                                                  diethylamino-1-                                                               methylbutyl)amino]-                                                           2-methoxypyrido                                                               [3,2-b]quinoline                                                              dihydrochloride                                                               + vaccine                                                                     Non-Immunized,                                                                            None                                                              22-21-20    14/30                                                                              2.31                                                         7-Chloro-10-[(4-                                                              diethylamino-1-                                                               methylbutyl)amino]-                                                           2-methoxypyrido                                                               [3,2-b]quinoline                                                              dihydrochloride,                                                              Treated                                                                       Non-Immunized,                                                                            None None   13/30        1.84.sup.4                               Infected Controls                                                             __________________________________________________________________________     .sup.1 Influenza A/Port Chalmers, solventextracted vaccine  30 CCA/mouse      by subcutaneous route.                                                        .sup.2 Compound given by gavage at 300 mg/kg at indicated time.               .sup.3 Significantly different from vaccine control group at P <0.05 on       basis of historical data.                                                     .sup.4 Ratio of lung weight/body weight for noninfected, nontreated, age      conditioned controls is characteristically less than 1.0 (0.65-0.80).    

                                      TABLE IV                                    __________________________________________________________________________    "Adjuvant" Effect on the Immunogenicity of                                    Influenzavirus A Vaccine                                                                  Day of Treatment                                                   Treatment Group                                                                           Relative to Infection Vaccine.sup.1 Drug.sup.2                                           Survivors/Total (14 Days Post-Infection)                                                   ##STR4##                                 __________________________________________________________________________    Vaccine Control                                                               21               None    8/10       1.21                                      7-Chloro-10-[(7-di-                                                           21                                                                            22-21-20    9/9  0.99.sup.3                                                   ethylaminoheptyl)                                                             amino]-2-methoxy-                                                             pyrido [3,2-b]qui-                                                            noline dihydrochlor-                                                          ide + vaccine                                                                 7-Chloro-10-[(6-di-                                                           21                                                                            22-21-20    10/10                                                                              0.88.sup.3                                                   methylaminohexyl)                                                             amino]-2-methoxy-                                                             pyrido[3,2-b]quinoline                                                        dihydrochloride + vac-                                                        cine                                                                          Non-immunized,                                                                            None None    4/10       1.83.sup.4                                Infected Control                                                              __________________________________________________________________________     .sup.1 Influenza A/Port Chalmers, solventextracted vaccine  30 CCA/mouse      by subcutaneous route.                                                        .sup.2 Compounds given by gavage at 300 mg/kg at indicated time.              .sup.3 Significantly different from vaccine control group at P <0.05 on       basis of historical data.                                                     .sup.4 Ratio of lung weight/body weight for noninfected, nontreated, age      conditioned controls is characteristically less than 1.0 (0.65-0.80).    

                                      TABLE V                                     __________________________________________________________________________    "Adjuvant" Effect on the Immunogenicity of Influenzavirus A Vaccine                             Day of Treatment                                             Treatment Group   Relative to Infection Vaccine.sup.1 Drug.sup.2                                          Survivors/Total (14 Days Post-Infection)                                                     ##STR5##                          __________________________________________________________________________    Vaccine Control                                                               21                     None  10/10         1.33                               7-Chloro-10-[5-di-                                                            21                                                                            22-21-20          10/10                                                                              1.12.sup.3                                             ethylaminopentyl)                                                             amino]-2-methoxypy-                                                           rido[3,2-b]quinoline                                                          dihydrochloride + vac-                                                        cine                                                                          2,2'-[[3-[(7-Chloro-                                                          21                                                                            22-21-20           9/10                                                                              1.03.sup.3                                             2-methoxypyrido[3,2-b]                                                        quinolin-10-yl)amino]                                                         propyl]imino]diethanol                                                        dihydrochloride + vaccine                                                     7-Chloro-10-[(4-di-                                                           21                                                                            22-21-20          10/10                                                                              0.97.sup.3                                             ethylamino-1-methyl-                                                          butyl)amino]-2-methoxy-                                                       pyrido[3,2-b]quinoline                                                        dihydrochloride + vaccine                                                     Non-Immunized, 7- None                                                        22-21-20           7/10                                                                              1.76                                                   Chloro-10-[(4-diethyl-                                                        amino-1-methylbutyl)amino]-                                                   2-methoxypyrido[3,2-b]qui-                                                    noline dihydrochloride, Treated                                               Non-Immunized,    None None   6/10         2.41.sup.4                         Infected Controls                                                             __________________________________________________________________________     .sup.1 Influenza A/Port Chalmers, solventextracted vaccine  30 CCA/mouse      by subcutaneous route.                                                        .sup.2 Compounds given by gavage at 300 mg/kg at indicated time.              .sup.3 Significantly different from vaccine control group at P <0.05 on       basis of historical data.                                                     .sup.4 Ratio of lung weight/body weight for noninfected, nontreated, age      conditioned controls is characteristically less than 1.0 (0.65-0.80).    

The compounds of the present invention are active orally asimmunostimulants. A range of doses may be employed depending on the modeof administration. For oral administration, these compounds are usuallyadministered at from about 37.5 to about 600 mg./kg./day.

In therapeutic use, the compounds of this invention may be administeredin the form of conventional pharmaceutical compositions. Suchcompositions may be formulated so as to be suitable for oraladministration. The active ingredient may be combined in admixture witha pharmaceutically acceptable carrier, which carrier may take a widevariety of forms, depending on the form or preparation desired foradministration. These compounds can be used on compositions such astablets. Here, the principal active ingredient is mixed withconventional tabletting ingredients such as corn starch, lactose,sucrose, sorbitol, talc, stearic acid, magnesium stearate, dicalciumphosphate, gums or similar materials such as non-toxic pharmaceuticallyacceptable diluents or carriers. The tablets or pills of these novelcompositions can be laminated or otherwise compounded to provide adosage from affording the advantage of prolonged or delayed action orpredetermined successive action of the enclosed medication. For example,the tablet or pill can comprise an inner dosage and an outer dosagecomponent, the latter being in the form of an envelope over the former.The two components can be separated by an enteric layer which serves toresist disintegration in the stomach and permits the inner component topass intact into the duodenum or to be delayed in release. A variety ofmaterials can be used for such enteric layers or coatings, suchmaterials including a number of polymeric acids or mixtures of polymericacids with such materials as shellac, shellac and cetyl alcohol,cellulose acetate and the like. A particularly advantageous entericcoating comprises a styrene-maleic acid copolymer together with knownmaterials contributing to the enteric properties of the coating. Thetablet or pill may be colored through the use of an appropriatenon-toxic dye.

The liquid forms in which the novel compositions of the presentinvention may be incorporated for administration include suitableflavored suspensions, elixirs, emulsions, solutions and similarpharmaceutical vehicles.

These dosage forms refer to physically discrete units suitable asunitary dosage for warm-blooded animals, each unit containing apredetermined quantity of active component calculated to produce thedesired therapeutic effect in association with the requiredpharmaceutical diluent, carrier or vehicle. The specification for thenovel dosage forms of this invention are indicated by characteristics ofthe active component. Examples of suitable dosage forms are tablets,capsules, pills, powder packets, granules, wafers, cachets,teaspoonfuls, dropperfuls, segregated multiples of any of the foregoingand other forms as herein described.

EXAMPLE 17-Chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

This compound may be prepared as described in Chemical Abstracts, 45,8531b (1951) or as described by Besley, D. M. and Goldberg, A. A. in J.Chem. Soc., 2448-2455, (1954) or it may be purchased from AldrichChemical Co.

EXAMPLE 27-Chloro-10-[(4-diethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

A mixture of 2.8 g. of 7,10-dichloro-2-methoxypyrido[3,2-b]quinoline,1.5 g. of 4-diethylaminobutylamine and 6.5 g. of phenol is heated at100° C. for 3 hours. The reaction is cooled, 20 ml. of ethanol is addedand this mixture is poured into a mixture of 200 ml. of acetone and 2.5ml. of concentrated hydrochloric acid. The yellow solic is collected,air-dried and recrystallized from ethanol, giving the desired product,mp. 246°-274° C. (dec.).

EXAMPLE 37-Chloro-10-[(4-dimethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

A mixture of 4.2 g. of 7,10-dichloro-2-methoxypyrido [3,2-b]quinoline,1.6 g. of dimethylaminobutylamine and 10 g. of phenol is stirred andheated on a steam bath for 3 hours. The reaction is cooled, 30 ml. ofethanol is added and this mixture is poured into a mixture of 300 ml. ofacetone and 3.75 ml. of concentrated hydrochloric acid. The yellowprecipitate is collected, washed with acetone, air-dried andrecrystallized from 100 ml. of 95% alcohol giving the desired product asyellow crystals, mp. 237°-238° C. (dec.)

EXAMPLE 47-Chloro-10-[(7-diethylaminoheptyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

A mixture of 2.8 g. of 7,10-dichloro-2-methoxypyrido [3,2-b]quinoline,2.0 g. of 7-diethylaminoheptylamine and 6.5 g. of phenol is stirred on asteam bath for 3 hours. The reaction is cooled, 20 ml. of 95% alcohol isadded and this mixture is poured into a mixture of 200 ml. of acetoneand 2.5 ml. of concentrated hydrochloric acid. The resulting solution iscooled in ice. The solid is collected and recrystallized fromn-propanol:ethyl acetate (50:50), giving the desired product as yellowcrystals, mp. 170°-174° C. (dec.).

EXAMPLE 57-Chloro-10-[(6-dimethylaminohexyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

A mixture of 2.8 g. of 7,10-dichloro-2-methoxypyrido [3,2-b]quinoline,1.5 g. of dimethylaminohexylamine and 6.5 g. of phenol is stirred andheated on a steam bath for 3 hours. The mixture is cooled, 20 ml. ofethanol are added and the dark solution is poured into a mixture of 200ml. of acetone and 2.5 ml. of concentrated hydrochloric acid. Themixture is cooled in ice. The solid is collected, air-dried andrecrystallized from 3A alcohol:acetone (50:50), giving the desiredproduct as yellow crystals, mp. 243°-244° C. (dec.).

EXAMPLE 68-Chloro-10-[(4-dimethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

To a solution of 19.1 g. of 2,5-dichlorobenzoic acid [Besley & Goldberg,J. Chem. Soc., 2448 (1954)] in 100 ml. of amyl alcohol is added 6.9 g.of anhydrous potassium carbonate and 0.1 g. of copper oxide. The mixtureis stirred and heated at 100° C. for 30 minutes, then 13.0 g. of5-amino-2-methoxypyridine is added and the mixture is heated at refluxand stirred for 3 hours. A small amount of water forms and is collectedin a Dean-Stark apparatus. The reaction mixture is cooled, poured intowater and made basic with potassium carbonate. The mixture issteam-distilled to remove the amyl alcohol, then filtered and thefiltrate is acidified to pH 6. The mixture is again filtered and thispink filtrate is acidified with 6 N hydrochloric acid to pH 4. The solidis collected, giving 5-chloro-2-(6-methoxy-3-pyridylamino)benzoic acidas a pink crystalline solid.

A mixture of 8.4 g. of the above product and 60 ml. of phosphorylchloride is heated under reflux for 51/2 hours. The excess phosphorylchloride is removed in vacuo and the solid residue is broken up in amixture of 250 g. of ice and 70 ml. of concentrated ammonium hydroxide.The mixture is maintained in an ice bath, with stirring, for 6 hours.The solid is collected, slurried in acetone, filtered, dried andrecrystallized from boiling dimethylformamide, giving8,10-dichloro-2-methoxybenzo[b]-1,5-naphthyridine as pale greencrystals.

A mixture of 2.8 g. of the preceding compound, 1.3 g. ofdimethylaminobutylamine and 6.5 g. of phenol is stirred and heated on asteam bath for 4 hours. The reaction is cooled, 20 ml. of 3A alcohol isadded and this solution is poured into a solution of 2.5 ml. ofconcentrated hydrochloric acid and 200 ml. of acetone. The solid iscollected and recrystallized from 3A alcohol, giving the desired productas yellow crystals, mp. 251°-252° C. (dec.)

EXAMPLE 77-Chloro-10-[(5-diethylaminopentyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride

A mixture of 2.8 g. of 7,10-dichloro-2-methoxypyrido [3,2-b]quinoline,1.7 g. of 5-diethylaminopentylamine and 6.5 g. of phenol was treated asin Example 5 to obtain the title compound, m.p. 232°-233° C.

EXAMPLE 82,2'-[[3-[(7-Chloro-2-methoxypyrido[3,2-b]quinolin-10-yl)amino]propyl]imino]diethanoldihydrochloride

A mixture of 2.8 g. of 7,10-dichloro-2-methoxypyrido [3,2-b]quinoline,1.8 g. of 3-[bis(2-hydroxyethyl)amino]propylamine and 6.5 g. of phenolwas treated as in Example 5 to obtain the title compound, m.p. 243°-245°C.

EXAMPLE 9 Preparation of Compressed Tablet

    ______________________________________                                        Ingredient              mg./Tablet                                            ______________________________________                                        Active compound         5-500                                                 Dibasic calcium phosphate N. F.                                                                       qs                                                    Starch U.S.P.           40                                                    Modified starch         10                                                    Magnesium stearate U.S.P.                                                                             1-5                                                   ______________________________________                                    

EXAMPLE 10 Preparation of Hard Shell Capsule

    ______________________________________                                        Ingredient            mg./Capsule                                             ______________________________________                                        Active compound       5-500                                                   Lactose, spray dried  qs                                                      Magnesium stearate    1-10                                                    ______________________________________                                    

EXAMPLE 11 Preparation of Oral Liquid

    ______________________________________                                        Ingredient             % W/V                                                  ______________________________________                                        Active compound        0.5-5                                                  Liquid sugar           75.0                                                   Methyl paraben U.S.P.   0.18                                                  Propyl paraben U.S.P.   0.02                                                  Flavoring agent        qs                                                     Purified water    qs ad                                                                              100.0                                                  ______________________________________                                    

We claim:
 1. A method of stimulating the immune response in awarm-blooded animal which comprises orally administering to said animalan effective immunostimulating amount of a compound of the formula:##STR6## wherein R₁ and R₂ are each selected from the group consistingof hydrogen and halogen; A is a straight or branched alkyl chain of 3 to7 carbon atoms; B is selected from the group consisting of dimethyl,diethyl and di-(2-hydroxyethyl); and the pharmaceutically acceptablesalts thereof.
 2. A method according to claim 1, wherein the compound is7-chloro-10-[(4-diethylamino-1-methylbutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride.
 3. A method according to claim 1, wherein the compoundis7-chloro-10-[(4-diethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride.
 4. A method according to claim 1, wherein the compoundis 7-chloro-10-[(4-dimethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinoline dihydrochloride.
 5. A method according to claim 1,wherein the compound is7-chloro-10-[(7-diethylaminoheptyl)amino]-2-methoxypyrido[3,2-b]quinoline dihydrochloride.
 6. A method according to claim 1,wherein the compound is7-chloro-10-[(6-dimethylaminohexyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride.
 7. A method according to claim 1, wherein the compoundis8-chloro-10-[(4-dimethylaminobutyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride.
 8. A method according to claim 1, wherein the compoundis7-chloro-10-[(5-diethylaminopentyl)amino]-2-methoxypyrido[3,2-b]quinolinedihydrochloride.
 9. A method according to claim 1, wherein the compoundis2,2'-[[3-[(7-chloro-2-methoxypyrido[3,2-b]quinolin-10-yl)amino]propyl]imino]diethanoldihydrochloride.